Use of cotinine in treating psychiatric disorders

ABSTRACT

A pharmaceutical composition is provided that is useful in treating obsessive-compulsive disorder, Tourette&#39;s Syndrome and schizophrenia comprising an amount of cotinine or a pharmaceutically acceptable salt thereof, which amount is effective to reduce or alleviate at least one of the symptoms of Tourette&#39;s Syndrome, obsessive-compulsive disorder, or schizophrenia in a human or other mammal.

FIELD OF THE INVENTION

This invention relates to treatment of certain psychological disordersthrough the administration of cotinine or salts of cotinine.

BACKGROUND OF THE INVENTION

U.S. Pat. No. 3,870,794 describes the use of cotinine in reducing anger,hostility, irritability and frustration, fear and anxiety withoutgeneral sedation effects. U.S. Pat. No. 5,187,169 describes theeffectiveness of cotinine in treating Alzheimer's Disease andParkinson's disease through its ability to bind to and hence causeactivation of nicotinic cholinergic receptors of the brain of thepatient while acting as a nicotinic agonist. Australian Pat. No. 273715describes the use of cotinine as an effective tranquilizer through itsaction as a muscle relaxant or anti-spasmodic and in lowering bloodpressure. Cotinine has also been shown to assist in tobacco withdrawal(pending U.S. application Ser. No. 124,004 and 885,314) as well as inhuman body weight management (pending U.S. application Ser. No. 964,277)and as a therapeutic agent in treating inflammatory bowel disorder suchas Crohn's disease or ulcerative colitis in humans (pending U.S.application Ser. No. 08/405,607).

It has not, however, been previously recognized that cotinine is alsoable to provide a therapeutic effect in psychiatric disorders such asobsessive-compulsive behavior, Tourette's Syndrome and schizophreniawhich are characterized by irrational behavior or repetitive thoughts.In obsessive-compulsive behavior, the patient is under a compulsion torepeat irrational or inappropriate behavior patterns and/or thoughts,while in Tourette's Syndrome the patient has an uncontrollablecompulsion to utter inappropriate sounds or words. Similarly, inschizophrenia, the patient has uncontrolled inappropriate or irrationalthoughts, delusions or behavior patterns. Schizophrenia, of course, hasother characteristics, primarily disorganized or impoverished speech orbehavior, flattened affect and avolition, social withdrawal anddiminished interest in school or work. While a number of pharmacologicaltreatments have been used for these psychiatric disorders in recentyears with a degree of success, no studies have reported the successfultreatment of all subjects or the absence of unwanted side effects, e.g.,depression, drowsiness, sedation, tardive dyskinesia, agranulocytosis,and other problems. Consequently, a continuing need exists forpharmacological treatments that will alleviate or reduceobsessive-compulsive disorder, Tourette's Syndrome and schizophrenia.

These and other more detailed and specific objects of the presentinvention will be better understood by reference to the followingdetailed description which illustrates by way of example but a few ofthe various forms of the invention within the scope of the appendedclaims.

SUMMARY OF THE INVENTION

In accordance with the present invention, cotinine or thepharmaceutically acceptable salts thereof, such as the tartrate,aspartate, lactate, malate, citrate, fumarate, sulfate or chloridesalts, are used in treating obsessive-compulsive behavior, Tourette'sSyndrome, and schizophrenia. The present invention provides atherapeutic method of treatment to reduce or alleviate the symptoms ofthe aforesaid disorders. In a preferred embodiment, the presentinvention provides a therapeutic method to alleviate or reduce thesymptoms of obsessive-compulsive disorder, Tourette's Syndrome andschizophrenia through the administration of any suitable dosage form,e.g., oral administration, injection, sublingual absorption, as aconstituent of chewing gum to be chewed by the patient, as a transdermalpatch to be applied to the skin of the patient, or through othersuitable forms of administration.

The present invention also provides, as an article of manufacture, apackaging material such as a box, bottle, tube, spray or insufflator,intravenous bag, envelope or the like and at least one unit dosage formof a pharmaceutical agent contained in the package wherein thepharmaceutical agent comprises cotinine or a pharmaceutically acceptablesalt thereof in an amount effective to alleviate or reduce the symptomsof obsessive-compulsive disorder, Tourette's Syndrome or schizophrenia,and wherein the package includes instructions indicating use foralleviating obsessive-compulsive disorder, Tourette's Syndrome orschizophrenia. Suitable instructions include printed labels, printedpackage inserts, tags, cassette tapes and the like.

DETAILED DESCRIPTION OF THE INVENTION

This invention provides a new and useful medicinal treatment used inalleviating or reducing the symptoms of obsessive-compulsive disorder,Tourette's Syndrome and schizophrenia in humans or other mammals. Thetreatment is effective in altering the emotional state of the humansubject treated by the pharmaceutical agent. Both the behavior andemotional state, namely a compulsion by the patient to repeat thoughtsor exhibit inappropriate or irrational behavior, are alleviated orreduced without unwanted side effects such as depression, drowsiness orsedation. In addition, the inappropriate behavior and/or irrationalthoughts are reduced and supplanted by improved and more appropriatebehavior and thought processes.

The treatment concerns the administration to a mammal, and especially ahuman being, in a pharmaceutically acceptable dosage form, atherapeutically effective amount of cotinine or its pharmacologicallyacceptable salts for alleviating or reducing the symptoms ofobsessive-compulsive disorder, Tourette's Syndrome and schizophrenia.

It is known that after a person consumes nicotine, the nicotine ismetabolized in four to six hours to produce cotinine. Cotinine begins toaccumulate in the blood shortly after nicotine is consumed but, becauseof the toxicity of nicotine, cotinine is present in relatively lowconcentrations in the blood as the result of consuming nicotine.Nicotine has a half-life in the human of about one to two hours, whereascotinine has a half-life of about 16 to 17 hours, and consequently ispresent in the blood much longer than nicotine. A one-pack-a-day smokerwould usually have a nicotine blood concentration of about 20 ng/ml orabout 1/10 μmol/ to about 1 μmol/l, while a one-half-pack-a-day smokerwill have a cotinine level of about 300 ng/ml or about 2 μmol/l. Becausecotinine is virtually nontoxic, it can be present in the blood inrelatively high amounts, say, 18,000 ng/ml without producing deleteriouseffects in humans. In accordance with the present invention, cotinine isused in amounts which produce much higher blood levels than that foundin smokers.

Cotinine has been shown in U.S. Pat. No. 5,187,169 to pass theblood-brain barrier in the treatment of Alzheimer's and Parkinson'sdiseases, disorders that are correlated with a loss of cholinergicmediated neurotransmission. Because these diseases respond to cotinine,it is theorized that cotinine acts on cholinergic neurotransmission.There are, of course, many different types of cholinergic neuronreceptors in mammals, and cotinine may express different affinity forand produce different effects on these various receptors.

There is also evidence that the weight control utility of cotinine, aswell as the benefit in relieving muscle spasms and treatinganger/hostility, result from cotinine's ability to modulate directly orindirectly other neurotransmitters such as serotonin, norepinephrine,dopamine or glutamate (U.S. Pat. No. 5,187,169).

While the effectiveness of cotinine for treating obsessive-compulsivedisorder, Tourette's Syndrome and schizophrenia has never appeared inthe literature, the relationship between nicotine and schizophrenia hasbeen noted, e.g., through the recognition that almost all schizophrenicssmoke incessantly, apparently in an attempt at self-medication FreedmanR., Hall M., Adler L. E., Leonard S. "Evidence in Postmortem BrainTissue for Decreased Number of Hippocampal Nicotinic Receptors inSchizophrenia. Biol. Psychiatry. 38(1):22-33 (1995)!. It is also commonto see many participants in Alcoholics Anonymous or Narcotics Anonymousgroups smoking feverishly. Moreover, it has been recognized by theHazelden Treatment Center at Center City, Minn., that continued smokingis considered to be important for a patient's success in his or heranti-addiction program. From these and other observations together withthe discoveries reported hereinbelow, I find adequate evidence toconclude that these patients are trying to self-medicate their addictiveor psychiatric disorders by consuming nicotine. In other words, thepatients are reducing disease symptoms by consuming nicotine which isquickly metabolized to cotinine.

Nicotine has been shown to be effective in the treatment of Tourette'sSyndrome McConville B. J., Sanberg P. R., Fogelson M. H., King J.,Cirino P., Parker K. W., Norman A. B. "The Effects of Nicotine PlusHaloperidol Compared to Nicotine Only and Placebo Nicotine Only inReducing Tic Severity and Frequency in Tourette's Disorder." Biol.Psychiatry. 31(8):832-840 (1992)!. I have now found that data developedin the following Examples demonstrate that cotinine, by virtue of itslow toxicity and longer half-life, is even more effective in treatingTourette's Syndrome than is the administration of nicotine.

The effectiveness of nicotine in treating other psychological conditionshas been recognized in U.S. Pat. Nos. 5,187,169 and 5,298,257.Consequently, there is a clear relationship between nicotine and brainactivity. The role of the central nervous system nicotinic receptors inhuman cognitive function and the involvement of the central nicotinicreceptors in several degenerative brain disorders has been recognized byPaul Newhouse Newhouse P. A., Potter A., Corwin J., Lenox R."Age-related Effects of the Nicotinic Antagonist Mecamylamine onCognition and Behavior. Neuropsychopharmacology. 10(2):93-107 (1994)!.The effectiveness of cotinine itself has already been demonstrated inalleviating tobacco addiction. See, pending U.S. patent application Ser.Nos. 885,314 and 124,004. The present invention is based in part on thefinding described below in the examples that cotinine is the primarytherapeutic agent for these and other actions typically ascribed tonicotine. I have found, however, that since cotinine in contrast tonicotine is very nontoxic in humans, it is possible to alleviate orreduce the symptoms of obsessive-compulsive disorder, Tourette'sSyndrome and schizophrenia by providing massive doses of the drug asrequired to achieve therapeutic results so as to provide blood levelswhich cannot be tolerated with nicotine because of its toxicity.

Neuro-receptor binding site radioimmunoassay studies using human andother mammalian tissue described in the following examples with nicotineand cotinine demonstrate that cotinine at the same blood level is aseffective or more effective than nicotine in many receptor sites. Inaddition, any curative effect produced by nicotine inobsessive-compulsive disorder, Tourette's Syndrome and schizophrenia canbe achieved to a much greater degree with cotinine than with nicotine inaccordance with the findings of the present invention. The effectivenessof cotinine, which will be described in greater detail below, is shownfor example in human tissue neuro-receptor binding site studies to beabout 19 times as effective in binding as an equal concentration ofnicotine at the muscarinic-1 receptor and about five times as effectiveas an equal concentration of nicotine at the muscarinic-2 receptor whenbinding is expressed as percent inhibition of the receptor. Thus, forthese human receptors, cotinine is many times more effective thannicotine at the same concentration in inhibiting the receptor bindingactivity of those receptors and can be administered in much greateramounts.

In producing its therapeutic effect, cotinine may be acting as acholinergic agonist which is the effect usually attributed to nicotine.Thus, the cotinine acts essentially to slow down the over-activity ofcertain neural receptors, such as those that are receptors to dopamine,serotonin, acetylcholine, muscarine and the like. Cotinine's ability toalter neural communication mediated by the above neurotransmitters orthe combination of them is the apparent reason for its utility intreating obsessive-compulsive disorder, Tourette's Syndrome andschizophrenia.

The exact psychoactive neuroleptic activity of cotinine is unknown. Itis known, however, to have serotonergic activity. The present inventiondemonstrates several other actions of cotinine.

Tourette's Syndrome and Other Obsessive-Compulsive Disorders

Tourette's Syndrome is characterized by an autosomal dominant multipletic disorder with variable penetrance progressing to multiple complexmovements including respiratory and vocal tics, grunting, barking noisesevolving into compulsive utterances. The neuroleptic drug of choice ishaloperidol (HaIdol™). Clonidine and pimozide are also used. Haloperidolis a strong tranquilizer that is believed to act by interfering theaction of dopamine, thereby reducing anxiety and agitation and improvingcoherence. The effectiveness of nicotine in treating Tourette's Syndromewas described by B. J. McConville and P. R. Sanberg in BiologicalPsychiatry, 31(8):832-840 (1992). They demonstrated long-term effects ofnicotine from relatively short term application of nicotine treatmentwith and without concurrent neuroleptic therapy. In contrast to theprior art, Tourette's Syndrome is treated in accordance with the presentinvention by the direct administration of an effective amount ofcotinine or pharmacologically acceptable salts thereof for reducing oreliminating the symptoms of Tourette's Syndrome behavior. The Exampleswhich follow below indicate that treatment with cotinine alleviates thecompulsive speech behavior more effectively than nicotine. Besides beingmore effective than nicotine, cotinine lacks virtually any side effectsas well as having a relatively long terminal elimination half-life ofabout 16 hours versus two hours for nicotine, and consequently acts inreducing the symptoms of Tourette's Syndrome over a longer period oftime.

In two prior human studies, the pharmacokinetic profiles of intravenousand orally administered cotinine were examined without emphasis onmeasuring the subjective and/or physiological changes induced by thiscompound N.L. Benowitz et al., Clin. Pharm. and Ther. 34:604 (1983).P.J. DeSchepper et al., Eur. J Pharmacol. 31:583 (1987)!. Moreover,using an uncontrolled experimental design, Benowitz et al., Clin. Pharm.and Ther. 34:604 (1983), found that intravenous cotinine infusion over60 minutes produced no cardiovascular changes and significant decreasesin subjective ratings of desire to smoke, irritability, low energy andanxiety/tension. These changes were comparable to placebo-inducedchanges found in other experiments with nicotine. Using a rapid infusionof cotinine over 5 minutes, no significant changes in the subjectiveratings were observed. Consequently, Benowitz and his colleaguesconcluded that cotinine lacked significant pharmacologic activity inhumans. In spite of this, nicotine, the precursor of cotinine, has anestablished utility in treating Tourette's Syndrome B. J. McConville, P.R. Sanberg. Ibid., p.8!. I have now discovered that the directadministration of cotinine will produce greater binding activity thannicotine at many sites and is therefore indicated to be more effectivethan nicotine in the treatment of Tourette's Syndrome and can also beused in much higher blood concentrations. The effectiveness is believedto be primarily due my finding that cotinine mimics or exceeds theblocking effect of nicotine at many neural receptor sites.

A brief review of neural activity will provide more completeunderstanding of the invention. An electrical impulse traveling along anexcited neuron will trigger the release of neurotransmitters from theexcited cell into the synaptic junction at the end of the neuron,causing the next cell to take in or exude selected ions, e.g., calciumand sodium. By altering the flow of charges across the membrane of thesecond cell, the neurotransmitter chemical, e.g., acetylcholine, cangive rise to a new impulse. It is well known that nicotine couples witha part of the receptor molecule at the synapse designed to bind to theneurotransmitter acetylcholine. The nicotine thus bound to the receptorserves as an acetylcholine agonist mimicking the stimulatory effect ofthe naturally produced acetylcholine. It is now known that thenicotine-sensitive or nicotinic acetylcholine receptors in the synapticjunction are present in the central nervous system of all highervertebrates. The brain has many other receptors, including a class ofreceptors known as muscarinic receptors as well as numerous others whichwere tested in the examples that follow. Nicotine, however, has ablocking effect for other receptors in addition to thenicotinic-acetylcholine receptors, and it is the nicotinic-acetylcholinereceptors, as well as others, that are active in the chain of eventsthat leads to what is known as Tourette's Syndrome, a disorder which haspreviously been at least partially corrected by the administration ofnicotine. Sanberg (Ibid., p.8) has shown the effectiveness of treatingTourette's Syndrome with psychoactive (neuroleptic) agents such asHaldol™ which blocks dopamine receptors. He showed that nicotine makesHaldol™ more effective. He showed that nicotine with Haldol™ works evenbetter than Haldol™ alone, and that nicotine alone appears to beeffective in treating Tourette's Syndrome. Moreover, it was demonstratedthat the skin patches containing nicotine had a lasting benefit forweeks to months. The blocking effect of nicotine that was found at thereceptor sites is believed to be the same mechanism through whichcotinine is effective in treating Tourette's Syndrome. While theeffectiveness of the present invention is not known with certainty, itis speculated that new receptors synthesized by the body after exposureto cotinine may act essentially to cure the defect which causesTourette's Syndrome. An example of this is the likelihood that tyrosinehydroxylase activity is induced by cotinine at the receptor site.

Obsessive-Compulsive Disorder

Obsessive-compulsive disorder (OCD) is characterized by the presence ofrecurrent ideas and fantasies and repetitive impulses or actions thatthe patient recognizes as morbid and toward which he feels a stronginner resistance. Some research suggests that disturbances in thefunction of the basal ganglia, especially in the seratonin receptors maybe an important element in OCD. One established treatment of choice isthe administration of the neuroleptic drug clomipramine (Anafranil™).Research has indicated that clomipramine increases brain tissueconcentrations of certain nerve impulse transmitters, especiallyseratonin, by blocking seratonin re-uptake. Nicotine acts as an agonistat the neurotransmitter receptor site much in the same way as the drugclomipramine. Thus, clomipramine mimics the stimulatory effect of thenaturally produced acetylcholine so as to become bound to other receptorsites as well, and cotinine apparently acts in a similar manner asrevealed in the examples below to thereby alleviate or controlobsessive-compulsive behavior.

Schizophrenia

Schizophrenia is a disabling psychiatric disorder characterized bydelusions, hallucinations, disorganized or impoverished speech,obsessive or compulsive behavior, flattened affect, and avolition. Thetreatment of schizophrenia with drugs has been heavily influenced by thehypothesis that certain dopamine pathways are overactive Seeman P.,Niznik H. B. "Dopamine Receptors and Transporters in Parkinson's Diseaseand Schizophrenia." FASEB J. 4:2737-44 (1990)!. Evidence for dopamineover-activity in patients with schizophrenia includes the capacity ofanti-psychotic drugs to block dopamine receptors in vivo and in vitroCreese I., Burt D. R., Schneider S. H. "Dopamine Receptor BindingPredicts Clinical and Pharmacological Potencies of Anti-schizophrenicDrugs." Science. 192:481-3 (1976)!. In addition, clinical efficacy ofanti-psychotic drugs is, in general, highly correlated with theirability to block dopamine D₂ receptors Seeman P., Lee T., Chau-Wong M.,Wong K. "Anti-psychotic Drug Doses and Neuroleptic/Dopamine Receptors."Nature. 261:717-9 (1976)!. Moreover, dopamine agonists exacerbate thesymptoms and signs of schizophrenia. Other receptors may also beinvolved in the therapeutic or adverse effects of specific drugs;however, all effective anti-psychotic drugs block some type of dopaminereceptor Seeman P. "Dopamine Receptor Sequences: Therapeutic Levels ofNeuroleptics Occupy D9 Receptors, Clozapine Occupies D4."Neuropsychopharmacology, 7:261-84 (1992)!. One of the drugs, clozapine,that is highly effective in treating schizophrenia has a relatively highaffinity for dopamine D1 and D4, 5-HT₂ muscarine in alpha-adrenergicreceptors and is also a dopamine D2 receptor antagonist Seeman P.,Niznik, H. B. "Dopamine Receptors and Transporters in Parkinson'sDisease and Schizophrenia." FASEB J, 4:2737-44 (1990); Seeman P."Dopamine Receptor Sequences: Therapeutic Levels of Neuroleptics OccupyD2 Receptors, Clozapine Occupies D4." Neuropsychopharmacology. 7:261-84(1992)!.

Research conducted by Edward Levin at Duke University shows that a sideeffect resulting from haloperidol therapy in treating schizophreniacould be reduced or blunted with nicotine which, as explained above, isquickly converted to cotinine in the body. Cotinine is thereforeindicated to be of value in enhancing haloperidol therapy in thetreatment of schizophrenia. The effect of cotinine on glutamic aciddecarboxylase (GAD) described below should play an important role inenhancing haloperidol therapy in schizophrenia.

The in vitro tests using human and animal tissue that were conducted indeveloping the present invention show that cotinine has the same highaffinity for many of the same receptor sites as clozapine. Consequently,its action and effectiveness in schizophrenia is believed to be of asimilar origin.

Freedman has shown nicotine to be effective in normalizing thepsycho-physiological defects of schizophrenia. It is well known thatschizophrenics are heavy smokers. This finding supports an explanationfor the consumption of nicotine by these patients as a self-administeredtherapy. Cholinergic neurons may be involved in schizophrenia. It hasbeen shown that bungarotoxin-sensitive cholinergic receptors in thehippocampus are involved in duplicating a second sonic responsecharacteristic of schizophrenia. Nicotine appears to be effective ininhibiting typical schizophrenic activity when used in combination withmecamylamine (MEC). The use of nicotine is not, however, acceptable as atherapy for schizophrenia because a high dose of nicotine (which can betoxic) is needed and the effect is short-lived. Thus, tachyphylaxisoccurs in short order Freedman R., Hall M., Adler L. E., Leonard S."Evidence in Postmortem Brain Tissue for Decreased Number of HippocampalNicotinic Receptors in Schizophrenia. Biol. Psychiatry. 38(l):22-33(1995)!.

In tests described below conducted in the development of the presentinvention, cotinine was found to have greater receptor binding activitythan nicotine at the same concentration at many of the same receptorsites examined and can also be administered to humans at a much higherconcentration. In contrast to nicotine, cotinine can be used at a muchhigher blood level and, due to its greater half-life, is effective forabout eight times as long as nicotine. In addition, I have found thatcotinine has greater activity than nicotine at many of the same receptorsites that bind nicotine which further facilitates its effectiveness.

Cotinine Dosage Forms

Cotinine (1-methyl-5-(3-pyridinyl)-2-pyrrolidionone) has the formulashown below: ##STR1##

The physiologically active form is the (-)-isomer, so as used herein,the term "cotinine" includes (-)-cotinine, or the racemic form,(±)-cotinine. The free base, depicted above, can be employed in thepractice of the invention, as can the pharmaceutically acceptable salts.These include the amine-acid addition salts of nontoxic organic acids orinorganic acids, such as the tartarate, fumarate ("scotine"), citrate,maleate, malate, hydrobromide, hydrochloride, sulfate, phosphate and thelike. For example, see F. Vaitekunas, J Amer. Chem. Soc. 79:149 (1957).E. R. Bowman et al. in J Pharmacol. and Exp. Ther. 135:306 (1962) reportthe preparation of (-)-cotinine free base from (-)-nicotine. Thepreparation and purification of (-)-cotinine fumarate is described by N.L. Benowitz et al., Clin. Pharmacol. Ther. 34:604 (1983).

Cotinine is the major metabolite of nicotine which accumulates in thebody as a result of nicotine exposure and has previously been believedto be pharmacologically inactive. For example, see N. L. Benowitz, "TheUse of Biologic Fluid Samples in Assessing Tobacco Smoke Consumption",in Measurement in the Analysis and Treatment of Smoking Behavior, J.Grabowski et al., eds., NIDA Research Monograph No. 48, U.S. DHHS, PHS,ADAMHA (1983). In contrast to nicotine, cotinine has a relatively longterminal elimination half-life (two versus sixteen hours, respectively).Due to this pharmacological characteristic, cotinine has become theprincipally used objective biochemical marker of nicotine exposure incigarette smoking and/or cessation-related research paradigms.

While cotinine is a well-known metabolite of nicotine and is routinelymeasured in many laboratories, no systematic investigation of thephysiological and subjective effects produced by intravenous cotinineadministration has been performed in humans. K. I. Yamamoto, et al.,International J. Neuropharmacol. 4:359 (1965) reported that intravenouscotinine produced increases only slightly in EEG activity and behavioralarousal in cats with only a slight decrease in blood pressure. Insquirrel monkeys, intramuscular cotinine injections increased rates ofresponding on fixed interval schedules of reinforcement over a widerange of doses M. E. Risner et al., J Pharmacol. and Exp. Ther. 234:113(1985); S. R. Goldberg et al., Psychopharmacology. 97:295 (1989)!. Thesefindings, taken together, suggest that cotinine is behaviorally active.However, the pharmacologic mechanism of action has yet to be determined.

In two recent human studies, the pharmacokinetic profiles of intravenousand orally administered cotinine were examined without emphasis onmeasuring the subjective and/or physiological changes induced by thiscompound N. L. Benowitz et al., Clin. Pharmacol. and Ther. 34:604(1983); P. J. DeSchepper et al., Eur. J Pharmacol. 31:583 (1987)!.Moreover, using an uncontrolled experimental design, Benowitz et al.,Clin. Pharmacol. and Ther. 34:604 (1983), found that intravenouscotinine infusion over 60 minutes produced no cardiovascular changes andsignificant decreases in subjective ratings of desire to smoke,irritability, low energy and anxiety/tension. These changes werecomparable to placebo-induced changes found in other experiments withnicotine. Using a rapid infusion of cotinine over 5 minutes, nosignificant changes in the subjective ratings were observed.Consequently, Benowitz and his colleagues concluded that cotinine lackedsignificant pharmacologic activity in humans.

Administration and Dosages

While it is possible that, for use in therapy, cotinine and/or its saltsmay be administered as the pure chemicals, as by inhalation of a finepowder via an insufflator, it is preferable to present the activeingredient as a pharmaceutical formulation. The invention thus furtherprovides a pharmaceutical formulation comprising cotinine and/or apharmaceutically acceptable salt thereof, together with one or morepharmaceutically acceptable carriers thereof and, optionally, othertherapeutic and/or prophylactic ingredients. The carrier(s) must be`acceptable` in the sense of being compatible with the other ingredientsof the formulation and not deleterious to the recipient thereof.

Pharmaceutical formulations include those suitable for oral orparenteral (including intramuscular, subcutaneous and intravenous)administration. Forms suitable for parenteral administration alsoinclude forms suitable for administration by inhalation or insufflationor for nasal, or topical (including buccal, rectal, vaginal andsublingual) administration. The formulations may, where appropriate, beconveniently presented in discrete unit dosage forms and may be preparedby any of the methods well known in the art of pharmacy. Such methodsinclude the step of bringing into association the active compound withliquid carriers, solid matrices, semi-solid carriers, finely dividedsolid carriers or combinations thereof, and then, if necessary, shapingthe product into the desired delivery system.

Pharmaceutical formulations suitable for oral administration may bepresented in discrete unit dosage forms such as hard or soft gelatincapsules, cachets or tablets each containing a predetermined amount ofthe active ingredient; as a powder or as granules; as a solution, asuspension or as an emulsion; in a chewable base such as a syntheticresin or chicle for ingestion of the cotinine from chewing gum. Theactive ingredient may also be presented as a bolus, electuary or paste.Tablets and capsules for oral administration may contain conventionalexcipients such as binding agents, fillers, lubricants, disintegrants,or wetting agents. The tablets may be coated according to methods wellknown in the art, i.e., with enteric coatings.

Oral liquid preparations may be in the form of, for example, aqueous oroily suspensions, solutions, emulsions, syrups or elixirs, or may bepresented as a dry product for constitution with water or other suitablevehicle before use. Such liquid preparations may contain conventionaladditives such as suspending agents, emulsifying agents, non-aqueousvehicles (which may include edible oils), or preservatives.

The compounds according to the present invention may also be formulatedfor parenteral administration (e.g., by injection, for example, bolusinjection or continuous infusion) and may be presented in unit dosageforms in ampules, pre-filled syringes, small volume infusion containersor multi-dose containers with an added preservative. The compositionsmay take such forms as suspensions, solutions, or emulsions in oily oraqueous vehicles, and may contain formulatory agents such as suspending,stabilizing and/or dispersing agents. Alternatively the activeingredient may be in powder form, obtained by aseptic isolation ofsterile solid or by lyophilization from solution, for constitution witha suitable vehicle, e.g., sterile, pyrogen-free water, before use.

For topical administration to the epidermis, the cotinine may beformulated as ointments, creams or lotions, or as the active ingredientof a transdermal patch. Suitable transdermal delivery systems aredisclosed, for example, in A. Fisher et al. (U.S. Pat. No. 4,788,603) orR. Bawa et al. (U.S. Pat. Nos. 4,931,279; 4,668,506; and 4,713,224).Ointments and creams may, for example, be formulated with an aqueous oroily base with the addition of suitable thickening and/or gellingagents. Lotions may be formulated with an aqueous or oily base and willin general also contain one or more emulsifying agents, stabilizingagents, dispersing agents, suspending agents, thickening agents, orcoloring agents. The active ingredient can also be delivered viaiontophoresis, e.g., as disclosed in U.S. Pat. Nos. 4,140,122;4,383,529; or 4,051,842.

Formulations suitable for topical administration in the mouth includeunit dosage forms such as lozenges comprising active ingredient in aflavored base, usually sucrose and acacia or tragacanth; pastillescomprising the active ingredient in an inert base such as gelatin andglycerin or sucrose and acacia; mucoadherent gels, and mouthwashescomprising the active ingredient in a suitable liquid carrier.

When desired, the above-described formulations can be adapted to givesustained release of the active ingredient employed, e.g., bycombination with certain hydrophilic polymer matrices, e.g., comprisingnatural gels, synthetic polymer gels or mixtures thereof.

Pharmaceutical formulations suitable for rectal administration whereinthe carrier is a solid are most preferably presented as unit dosesuppositories. Suitable carriers include cocoa butter and othermaterials commonly used in the art, and the suppositories may beconveniently formed by admixture of the active compound with thesoftened or melted carrier(s) followed by chilling and shaping in molds.

Formulations suitable for vaginal administration may be presented aspessaries, tampons, creams, gels, pastes, foams or sprays containing, inaddition to the active ingredient, such carriers as are known in the artto be appropriate.

For administration by inhalation, the compounds according to theinvention are conveniently delivered from an inhaler such as aninsufflator, nebulizer or a pressurized pack or other convenient meansof delivering an aerosol spray, smoke, vapor, mist or powder.Pressurized packs may comprise a suitable propellant such asdichlorodifluoromethane, trichlorofluoromethane,dichlorotetrafluoromethane, carbon dioxide or other suitable gas. In thecase of a pressurized aerosol, the dosage unit may be determined byproviding a valve to deliver a metered amount.

Alternatively, for administration by inhalation or insufflation, thecompounds according to the present invention may take the form of a drypowder composition, for example, a powder mix of the compound and asuitable powder base such as lactose or starch. The powder combinationmay be presented in unit dosage form in, for example, capsules orcartridges or, e.g., gelatin or blister packs from which the powder maybe administered with the aid of an inhalator or insufflator.

For intra-nasal administration, the compounds of the invention may beadministered via a liquid spray, such as via a plastic bottle atomizer.Typical of these are the Mistometer® (Wintrop) and the Medihaler®(Riker).

For topical administration to the eye, the cotinine can be administeredas drops, gels (see, S. Chrai et al., U.S. Pat. No. 4,255,415), gums(see, S. L. Lin et al., U.S. Pat. No. 4,136,177) or via aprolonged-release ocular insert (see, A. S. Michaels, U.S. Pat. No.3,867,519 and H. M. Haddad et al., U.S. Pat. No. 3,870,791).

The pharmaceutical compositions according to the invention may alsocontain other adjuvants such as flavorings, colorings, antimicrobialagents, or preservatives.

It will be further appreciated that the amount of cotinine, or an activesalt or derivative thereof, required for use in treatment will vary notonly with the particular salt selected but also with the route ofadministration, the nature of the condition being treated, and the ageand condition of the patient and will be ultimately at the discretion ofthe attendant physician or clinician.

In general, however, a suitable dose will be in the range of from about0.5 mg/kg to about 100 mg/kg, e.g., from about 5 mg/kg to about 75 mg/kgof body weight per day, such as 3 mg to about 50 mg per kilogram of bodyweight of the recipient per day, preferably in the range of 6 mg/kg/dayto 90 mg/kg/day, most preferably in the range of 5 mg/kg/day to 60mg/kg/day, calculated as (-)-cotinine in the free base form.

The compound is conveniently administered in unit dosage form; forexample, containing 5 mg to 1000 mg, conveniently 10 mg to 750 mg, mostconveniently, 40-50 mg to 100-500 mg of active ingredient per unitdosage form.

Ideally, the active ingredient should be administered to achieve peakplasma concentrations of the active compound of from about 0.5 μM,preferably about 1 μM to 50 μM, most preferably about 2 FM to about 30μM. This may be achieved, for example, by the intravenous injection of a0.5% to 50% solution of the active ingredient, optionally in saline, ororally administered as a bolus containing about 1-100 mg, preferablyabout 25-95 mg of the active ingredient. Desirable blood levels may bemaintained by continuous infusion to provide about 0.01-5.0 mg/kg/hr orby intermittent infusions containing about 0.4-15 mg/kg of the activeingredient(s).

The desired dose may conveniently be presented in a single dose or asdivided doses administered at appropriate intervals, for example, astwo, three, four or more sub-doses per day. The sub-dose itself may befurther divided, e.g., into a number of discrete loosely spacedadministrations; such as multiple inhalations from an insufflator or byapplication of a plurality of drops into the eye.

The preferred dose of the compound is that amount that is effective totreat the psychological disorder from which the patient is suffering.The term "effective amount" or "effective dose" means that amountsufficient to pass across the blood-brain barrier so as to bind torelevant receptor sites in the central nervous system of the patient andto elicit neuropharmacological effects, e.g., by modifyingneurotransmitter activity so as to be effective in treating the diseaseby decreasing the symptoms of the particular psychological disorderbeing treated.

Cotinine has a long in vivo half-life of about 16 to 17 hours, completeoral bioavailability, minimal effect on the cardiovascular system, andhas not been reported to be harmful even at very high doses in manyspecies, including man. Also, because cotinine has no significant effecton the heart, it can be used in much higher concentrations thannicotine.

The invention will be further described by reference to the followingexamples.

EXAMPLES

Tissue cultures were prepared using standard laboratory practice for thefollowing tissues: human recombinant Ml clone, human recombinant M₂clone, guinea pig striatal membranes, rat cortical membranes, ratstriatal membranes, rat forebrain membranes, bovine hippocampalmembranes, guinea pig cerebellar membranes, rat kidney membranes,recombinant rat 5HT₇ clone, and rat cerebellar membranes.Radioimmunoassay studies were conducted to determine the bindingactivity of nicotine, cotinine and cotinine fumarate as described in theliterature references listed in the examples for the sites,neurotransmitters and receptors indicated.

    __________________________________________________________________________    Example 1.                                                                    Dopamine Reuptake Binding Assay                                               __________________________________________________________________________    Reference Compounds Ki(nM)                                                                  Bupropion                                                                          935.0                                                      Assay Characteristics                                                         K.sub.d (binding affinity):                                                                 28.0 nM                                                         B.sub.max (receptor number):                                                                113 fmol/mg tissue (wet weight)                                 Degree of Specific Binding:                                                                 70-80% (Non-specific binding determine using 10.sup.-4 M                      bupropion)                                                      Materials and Methods:                                                        Receptor Source:                                                                            Guinea pig striatal membranes                                   Radioligand:   .sup.3 H!WIN,35,428 (60-87 Ci/mmol)                                          Final ligand concentration -  1.0 nM!                           Reference Compound:                                                                         Bupropion                                                       Positive Control:                                                                           Bupropion                                                       Incubation Conditions:                                                                      Reactions are carried out in 50 nM TRIS-HCl (pH 7.4)                          containing                                                                    100 nM NaCl 25° C. for 2 hours. The reaction is                        terminated by rapid                                                           vacuum filtration onto glass fiber filters. Radioactivity                     trapped onto                                                                  the filters is determined and compared to control values in                   order to                                                                      ascertain any interactions of test compound with the                          dopamine                                                                      uptake site.                                                                  Literature references to receptor binding activity                            procedure:                                                                    Madras, et al. Cocaine Receptors Labeled by  .sup.3                           H!2Beta-                                                                      Carbomethoxy-3-Beta-(4-Fluorophenyl)topane. Mol.                              Pharmacol.                                                                    36: 518-524 (1989) with modifications.                                        Javitch, J. J., Blaustein, R. O., and Snyder, S. H.  .sup.3                   H!Mazindol                                                                    Binding Associated with Neuronal Dopamine and                                 Norepinephrine                                                                Uptake Sites. Mol. Pharmacol. 26: 35-44 (1984).                 Results:      Receptor Binding Activity Percent Inhibition Average (N =                     2)                                                                            Nicotine Cotinine                                                                            Cotinine                                                                           Cotinine.sub.2 Fumarate                                   10.sup.-6 M                                                                            10.sup.-4 M                                                                         10.sup.-6 M                                                                        10.sup.-6 M                                               9.2%     7.5%  14.2%                                                                              4.6%                                        __________________________________________________________________________    Example 2.                                                                    Gamma-Aminobutyric Acid (GABA) Uptake Binding Assay                           __________________________________________________________________________    Reference Compounds Ki(nM)                                                                  Nipecotic Acid                                                                       37,000                                                                 GABA   26,000                                                   Assay Characteristics                                                         K.sub.d (binding affinity):                                                                 18,000 nM                                                       B.sub.max (receptor number):                                                                780 fmol/mg tissue (wet weight)                                 Degree of Specific Binding:                                                                 90% (Non-specific binding determine using 1 nM Nipecotic                      Acid)                                                           Materials and Methods:                                                        Receptor Source:                                                                            Rat cortical membranes                                          Radioligand:   .sup.3 H!GABA (70-80 Ci/mmol)                                                Final Iigand concentration -  3.6 nM!                           Reference Compound:                                                                         Nipecotic acid                                                  Positive Control:                                                                           Nipecotic acid                                                  Incubation Conditions:                                                                      Reactions are carried out in KREBS-HEPES (pH 7.4) buffer                      at                                                                            37° C. for 3 minutes. The reaction is terminated by                    rapid vacuum                                                                  filtration onto glass fiber filters. Radioactivity trapped                    onto the                                                                      filters is determined and compared to control values in                       order to                                                                      ascertain any interactions of test compound with the GABA                     uptake                                                                        site.                                                                         Literature references to receptor binding activity                            procedure:                                                                    Karbon, E. W., Enna, S. J., and Ferkany, J. W. Biochemical                    and                                                                           Behavioral Studies Following Subchronic Administration of                     GABA                                                                          Uptake Inhibitors in Mice. Neuropharmacology. 30:                             1198-1192                                                                     (1991).                                                                       Falch, E., Hedegaard, A., et al. Comparative                                  Stereostructure -                                                             Activity Studies on GABA.sub.A and GABA.sub.B Receptor                        Sites and GABA                                                                Uptake using Rat Brain Membranes. Jrnl. Neurochem. 47(3):                     898-903 (1986).                                                 Results:      Receptor Binding Activity Percent Inhibition Average (N =                     2)                                                                            Nicotine Cotinine                                                                            Cotinine                                                                           Cotinine.sub.2 Fumarate                                   10.sup.-6 M                                                                            10.sup.-4 M                                                                         10.sup.-6 M                                                                        10.sup.-6 M                                               12.1%    7.4%  13.9%                                                                              15.8%                                       __________________________________________________________________________    Example 3.                                                                    Nicotinic (Neuronal) Binding Assay                                            __________________________________________________________________________    Reference Compounds Ki(nM)                                                                  Nicotine Sulfate                                                                      1.7                                                                   Hexamethonium                                                                         >10,000                                                               Atropine                                                                              >10,000                                                               Tubocurarine                                                                          >10,000                                                 Assay Characteristics                                                         K.sub.d (binding affinity):                                                                 2.7 nM                                                          B.sub.max (receptor number):                                                                118.4 fmol/mg protein                                           Degree of Specific Binding:                                                                 80% (Non-specific binding determine using 0.2 μM                           nicotine sulfate)                                               Materials and Methods:                                                        Receptor Source:                                                                            Rat cortical membranes                                          Radioligand:   .sup.3 H!N-methylcarbamylcholine iodide (70-87 Ci/mmol)                      Final ligand concentration -  2.5 nM!                           Reference Compound:                                                                         Nicotine sulfate                                                Positive Control:                                                                           Nicotine sulfate                                                Incubation Conditions:                                                                      Reactions are carried out in 50 mM TRIS-HCl (pH 7.4)                          containing                                                                    120 mM NaCl, 5.0 mM Kcl, 2.0 mM CaCl.sub.2, 1.0 mM                            MgCl.sub.2 and 3.0                                                            μM atropine sulfate at 4° C. for 60 minutes. The                    reaction is                                                                   terminated by rapid vacuum filtration onto glass fiber                        filters.                                                                      Radioactivity trapped onto the filters is determined and                      compared                                                                      to control values in order to ascertain any interactions of                   test                                                                          compound with the nicotinic binding site.                                     Literature references to receptor binding activity                            procedure:                                                                    Boska, P. and Quirion, R.  .sup.3 H!N-Methyl-Carbamylcholine                  : A New                                                                       Radioligand Specific for Nicotinic Acetylcholine Receptors                    in the                                                                        Brain. Eur. Jrnl. Pharmacol. 139: 323-333 (1987) with                         modifications.                                                                Leutje, C. W., Patrick, J., and Seguela, P. Nicotine                          Receptors in                                                                  the Mammalian Brain. FASEB Jrnl. 4: 2753-2760 (1990).           Results:      Receptor Binding Activity Percent Inhibition Average (N =                     2)                                                                            Nicotine Cotinine                                                                            Cotinine                                                                           Cotinine.sub.2 Fumarate                                   10.sup.-6 M                                                                            10.sup.-4 M                                                                         10.sup.-6 M                                                                        10.sup.-6 M                                               98.5%    12.0% 78.5%                                                                              15.9%                                       __________________________________________________________________________    Example 4.                                                                    Dopamine (D.sub.1 Central) Binding Assay                                      __________________________________________________________________________    Reference Compounds Ki(nM)                                                                  SCH 23390                                                                             4.6                                                                   Butaclamol                                                                            37.3                                                                  Metoclopramide                                                                        97.3                                                                  SKF 38393                                                                             127.0                                                                 Spiperone                                                                             843.0                                                   Assay Characteristics                                                         K.sub.d (binding affinity):                                                                 5.3 nM                                                          B.sub.max (receptor number):                                                                69 fmol/mg tissue (wet weight)                                  Degree of Specific Binding:                                                                 90% (Non-specific binding determined using 1.0 μM SCH                      23390)                                                          Materials and Methods:                                                        Receptor Source:                                                                            Rat striata membranes                                           Radioligand:   .sup.3 H!SCH 23390 (70-87 Ci/mmol)                                           Final ligand concentration -  0.5 nM!                           Reference Compound:                                                                         SCH 23390                                                       Positive Control:                                                                           SCH 23390                                                       Incubation Conditions:                                                                      Reactions are carried out in 50 mM HEPES (pH 7.4)                             containing 1.0                                                                mM EDTA, 4.0 mM MgSO.sub.4, and 10 μM ketanserin at                        37° C. for 60                                                          minutes. The reaction is terminated by rapid vacuum                           filtration onto                                                               glass fiber filters. Radioactivity trapped onto the filters                   is                                                                            determined and compared to control values in order to                         ascertain                                                                     any interactions of test compound with the dopamine binding                   site.                                                                         Literature references to receptor binding activity                            procedure:                                                                    Billard, W., Ruperto, V., Crosby, G., et al.                                  Characterization of the                                                       Binding of  .sup.3 H!SCH 23390: A Selective D.sub.1                           Receptor Antagonist                                                           Ligand in Rat Striatum. Life Sciences. 35: 1885-1893 (1984)                   with                                                                          modifications.                                                                Anderson, P. H., Gronvald, F. C., et al. NNC-112, NNC-687,                    and                                                                           NNC-756, New Selective and Highly Potent Dopamine D.sub.1                     Receptor                                                                      Antagonists. Eur. Jrnl. Pharma. 219(1): 45-52 (1992).           Results:      Receptor Binding Activity Percent Inhibition Average (N =                     2)                                                                            Nicotine Cotinine                                                                            Cotinine                                                                           Cotinine.sub.2 Fumarate                                   10.sup.-6 M                                                                            10.sup.-4 M                                                                         10.sup.-6 M                                                                        10.sup.-6 M                                               2.6%     3.3%  2.9% -1.2%                                       __________________________________________________________________________    Example 5.                                                                    Histamine (H.sub.2) Binding Assay                                             __________________________________________________________________________    Reference Compounds Ki(nM)                                                                  Cimetidine                                                                          650                                                       Assay Characteristics                                                         K.sub.d (binding affinity):                                                                 9.4 μM                                                       B.sub.max (receptor number):                                                                212 fmol/mg protein                                             Degree of Specific Binding:                                                                 60% (Non-specific binding determined using 10 mM                              cimetidine)                                                     Materials and Methods:                                                        Receptor Source:                                                                            Guinea pig striatal membranes                                   Radioligand:   .sup.3 H!Tiotidine (70-90 Ci/mmol)                                           Final ligand concentration -  4.0 nM!                           Reference Compound:                                                                         Cimetidine                                                      Positive Control:                                                                           Cimetidine                                                      Incubation Conditions:                                                                      Reactions are carried out in 50 mM Na--KPO.sub.4 (pH 7.4)                     at 25° C. for                                                          20 minutes. The reaction is terminated by rapid vacuum                        filtration                                                                    onto glass fiber filters. Radioactivity trapped onto the                      filters is                                                                    determined and compared to control values in order to                         ascertain                                                                     any interactions of test compound with the histamine.sub.2                    binding site.                                                                 Literature references to receptor binding activity                            procedure:                                                                    Gajtkowski, et al. Specific Binding of  .sup.3 H!Tiotidine                    to Histamine H.sub.2                                                          Receptors in Guinea Pig Cerebral Cortex. Nature. 304:                         65-67                                                                         (1983) with modifications.                                                    Martinez-Mur, M. I., Pollard, H., Moreau, J., et al. Three                    Histamine                                                                     Receptors (H.sub.1, H.sub.2, and H.sub.3) Visualized in the                   Brain of Human and                                                            Non-Human Primates. Brain Res. 526: 322-327 (1990).                           Haaksma, E. E. J., Leurs, R., and Timmerman, H. Histamine                     Receptors: Subclasses and Specific Ligands. Pharmac. Ther.                    47:                                                                           73-104 (1990).                                                  Results:      Receptor Binding Activity Percent Inhibition Average (N =                     2)                                                                            Nicotine Cotinine                                                                            Cotinine                                                                           Cotinine.sub.2 Fumarate                                   10.sup.-6 M                                                                            10.sup.-4 M                                                                         10.sup.-6 M                                                                        10.sup.-6 M                                               45.9%    -1.4% 9.2% 14.7%                                       __________________________________________________________________________    Example 6.                                                                    Histamine (H.sub.3) Binding Assay                                             __________________________________________________________________________    Reference Compounds Ki(nM)                                                                  R(-)-α-Methylhistamine                                                             0.79                                                               Histamine  59.3                                                 Assay Characteristics                                                         K.sub.d (binding affinity):                                                                 0.37 nM                                                         B.sub.max (receptor number):                                                                73 fmol/mg protein                                              Degree of Specific Binding:                                                                 90% (Non-specific binding determined using 1.0 μM                          R(-)-α-Methylhistamine                                    Materials and Methods:                                                        Receptor Source:                                                                            Rat forebrain membranes                                         Radioligand:   .sup.3 H!N.sup.a -methylhistamine (84.1 Ci/mmol)                             Final ligand concentration -  0.2 nM!                           Reference Compound:                                                                         R(-)-α-Methylhistamine                                    Positive Control:                                                                           R(-)-α-Methylhistamine                                    Incubation Conditions:                                                                      Reactions are carried out in 50 mM TRIS-HCl (pH 7.5) for                      40                                                                            minutes at 25° C. The reaction is terminated by                        rapid vacuum                                                                  filtration onto glass flber filters. Radioactivity trapped                    onto the                                                                      filters is determined and compared to control values in                       order to                                                                      ascertain any interactions of test compound with the                          histamine.sub.3                                                               binding site.                                                                 Literature references to receptor binding activity                            procedure:                                                                    West, Robert E., et al. Identification of Two H.sub.3                         -Histamine Receptor                                                           Subtypes. Mol. Pharmacol. 38: 610-613 (1990) with                             modifications.                                                                Arrang, J. M., Garbarg, M., Lancelot, J. C., et al. Highly                    Potent and                                                                    Selective Ligands for Histamine H.sub.3 Receptors. Nature.                    327: 117-                                                                     123 (1987) with modifications.                                  Results:      Receptor Binding Activity Percent Inhibition Average (N =                     2)                                                                            Nicotine Cotinine                                                                            Cotinine                                                                           Cotinine.sub.2 Fumarate                                   10.sup.-6 M                                                                            10.sup.-4 M                                                                         10.sup.-6 M                                                                        10.sup.-6 M                                               13.5%    15.7% 11.9%                                                                              15.4%                                       __________________________________________________________________________    Example 7.                                                                    Serotonin (5HT.sub.1) Binding Assay                                           __________________________________________________________________________    Reference Compounds Ki(nM)                                                                  5-Carboxytryptamine (5-CT)                                                                 2.2                                                              5-Hydroxytryptamine (5-HT)                                                                 7.2                                                              5-Methoxytryptamine                                                                        45.8                                                             Methysergide 790.0                                                            CGS-12066B   1,055.0                                            Assay Characteristics                                                         K.sub.d (binding affinity):                                                                 2.8 nM                                                          B.sub.max (receptor number):                                                                9.2 fmol/mg protein                                             Degree of Specific Binding:                                                                 60% (Non-specific binding determined using 100 μM                          serotonin)                                                      Materials and Methods:                                                        Receptor Source:                                                                            Rat cortical membranes                                          Radioligand:   .sup.3 H!Hydroxytryptamine binoxalate (15-30 Ci/mmol)                        Final ligand concentration -  3.0 nM!                           Reference Compound:                                                                         Serotonin                                                       Positive Control:                                                                           Serotonin                                                       Incubation Conditions:                                                                      Reactions are carried out in 50 mM TRIS-HCl (pH 7.4) at                       37° C. for                                                             45 minutes. The reaction is terminated by rapid vacuum                        filtration                                                                    onto glass fiber filters. Radioactivity trapped onto the                      filters is                                                                    determined and compared to control values in order to                         ascertain                                                                     any interactions of test compound with the 5HT.sub.1                          binding site.                                                                 Literature references to receptor binding activity                            procedure:                                                                    Peroutka, S. J., Snyder, S. H. Multiple Serotonin                             Receptors:                                                                    Differential Binding of  .sup.3 H!-5-HT,  .sup.3 H!-LSD and                    .sup.3 H!-Spiroperidol.                                                      Mol. Pharmacol. 16: 687-699 (1979) with modifications.                        Peroutka, S. J. and Snyder, S. H. Two Distinct Serotonin                      Receptors:                                                                    Regional Variations in Receptor Binding in Mammalian                          Brain.                                                                        Brain Research. 208: 339-347 (1981).                                          Martin, G. R. and Humphrey, P. P. A. Classification Review                    for                                                                           5-HT: Current Perspectives on Classification and                              Nomenclature.                                                                 Neuropharmacol. 3(3/4): 261-273 (1994).                         Results:      Receptor Binding Activity Percent Inhibition Average (N =                     2)                                                                            Nicotine Cotinine                                                                            Cotinine                                                                           Cotinine.sub.2 Fumarate                                   10.sup.-6 M                                                                            10.sup.-4 M                                                                         10.sup.-6 M                                                                        10.sup.-6 M                                               -1.4%    -0.8% 10.9%                                                                              5.4%                                        __________________________________________________________________________    Example 8.                                                                    Serotonin (5HT.sub.1A) Binding Assay                                          __________________________________________________________________________    Reference Compounds Ki(nM)                                                                  8-OH-DPAT                                                                           3.6                                                                     RU 24969                                                                            10.0                                                                    Serotonin                                                                           12.4                                                                    Ketanserin                                                                          >10,000                                                   Assay Characteristics                                                         K.sub.d (binding affinity):                                                                 2.0 nM                                                          B.sub.max (receptor number):                                                                1.626 pmol/mg protein                                           Degree of Specific Binding:                                                                 90% (Non-specific binding determined using 10 μM                           serotonin)                                                      Materials and Methods:                                                        Receptor Source:                                                                            Bovine hippocampal membranes                                    Radioligand:   .sup.3 H!-8-OH-DPAT (100 Ci/mmol)                                            Final ligand concentration -  1.0 nM!                           Reference Compound:                                                                         8-OH-DPAT                                                       Positive Control:                                                                           8-OH-DPAT                                                       Incubation Conditions:                                                                      Reactions are carried out in 50 mM TRIS-HCl (pH 7.4) at                       37° C. for                                                             10 minutes. The reaction is terminated by rapid vacuum                        filtration                                                                    onto glass fiber filters. Radioactivity trapped onto the                      filters is                                                                    determined and compared to control values in order to                         ascertain                                                                     any interactions of test compound with the 5HT.sub.1A                         binding site.                                                                 Literature references to receptor binding activity                            procedure:                                                                    Hoyer, D. Engel, G., et al. Molecular Pharmacology of                         5HT.sub.1 and                                                                 5-HT.sub.2 Recognition Sites in Rat and Pig Brain                             Membranes:                                                                    Radioligand Binding Studies with  .sup.3 H!-5HT,  .sup.3                      H!8-OH-DPAT,                                                                   .sup.125 I!-Iodocyanopindolol,  .sup.3 H!-Mesulergine and                     .sup.3 H!-Ketanserin.                                                        Eur. Jrnl. Pharmacol. 118: 13-23 (1985) with                                  modifications.                                                                Schoeffter, P. and Hoyer, D. How Selective is GR 43175?                       Interactions with Functional 5-HT.sub.1A, 5-HT.sub.1B,                        5-HT.sub.1C, and 5-HT.sub.1D                                                  Receptors. Naunyn-Schmiedeberg's Arch. Pharmac. 340: 135-                     138 (1989) with modifications.                                  Results:      Receptor Binding Activity Percent Inhibition Average (N =                     2)                                                                            Nicotine Cotinine                                                                            Cotinine                                                                           Cotinine.sub.2 Fumarate                                   10.sup.-6 M                                                                            10.sup.-4 M                                                                         10.sup.-6 M                                                                        10.sup.-6 M                                               3.3%     1.4%  1.1% 2.7%                                        __________________________________________________________________________    Example 9.                                                                    Serotonin (Non-Selective) Binding Assay                                       __________________________________________________________________________    Reference Compounds ki(nM)                                                                  Methysergide                                                                            5.7                                                                 Spiroperidol                                                                            18.0                                                                Mianserin 33.0                                                                5-Methoxytryptamine                                                                     210.0                                                 Assay Characteristics                                                         K.sub.d (binding affinity):                                                                 7.2 nM                                                          B.sub.max (receptor number):                                                                23 fmol/mg tissue (wet weight)                                  Degree ofSpecific Binding:                                                                  75% (Non-specific binding determined using 1.0 μM LSD)       Materials and Methods:                                                        Receptor Source:                                                                            Rat cortical membranes                                          Radioligand:   .sup.3 H!Lysergic acid diethylamide (60-70 Ci/mmol)                          Final ligand concentration -  5.0 nM!                           Reference Compound:                                                                         Methysergide                                                    Positive Control:                                                                           Methysergide                                                    Incubation Conditions:                                                                      Reactions are carried out in 50 mM TRIS-HCl (pH 7.4)                          containing 4                                                                  mM CaCl.sub.2, 0.1 mM pargyline and 0.1% ascorbic acid at                     37° C. for                                                             15 minutes. The reaction is terminated by rapid vacuum                        flltration                                                                    onto glass fiber filters. Radioactivity trapped onto the                      filters is                                                                    determined and compared to control values in order to                         ascertain                                                                     any interactions of test compound with the serotonin                          binding site.                                                                 Literature references to receptor binding activity                            procedure:                                                                    Peroutka, S. J., Snyder, S. H. Multiple Serotonin                             Receptors:                                                                    Differential Binding of  .sup.3 H!-5-HT,  .sup.3 H!-LSD and                    .sup.3 H!-Spiroperidol.                                                      Mol. Pharmecol. 16: 687-699 (1979) with modifications.                        Peroutka, S. J. and Snyder, S. H. Two Distinct Serotonin                      Receptors:                                                                    Regional Variations in Receptor Binding in Mammalian                          Brain.                                                                        Brain Research. 208: 339-347 (1981).                            Results:      Receptor Binding Activity Percent Inhibition Average (N =                     2)                                                                            Nicotine Cotinine                                                                            Cotinine                                                                           Cotinine.sub.2 Fumarate                                   10.sup.-6 M                                                                            10.sup.-4 M                                                                         10.sup.-6 M                                                                        10.sup.-6 M                                               6.3%     12.1% 4.6% 4.1%                                        __________________________________________________________________________    Example 10.                                                                   Opiate (Delta) Binding Assay                                                  __________________________________________________________________________    Reference Compounds Ki(nM)                                                                  Naloxone                                                                            25.2                                                                    DADLE 0.9                                                                     DPDPE 1.6                                                                     Cyctazocine                                                                         4.8                                                                     Nalbuphine                                                                          124.0                                                                   Pentazocine                                                                         140.0                                                                   DAMGO 145.5                                                     Assay Characteristics                                                         K.sub.d (binding affinity):                                                                 2.12 nM                                                         B.sub.max (receptor number):                                                                1.03 fmol/mg tissue (wet weight)                                Degree of Specific Binding:                                                                 80% (Non-specific binding determined using 10 μM                           naloxone)                                                       Materials and Methods:                                                        Receptor Source:                                                                            Rat forebrain membranes                                         Radioligand:   .sup.3 H!Enkephaline  2-D penicillamine, 5-D                                 penicillamine!-(DPDPE)                                                        (30-60 Ci/mmol)                                                 Reference Compound:                                                                         Final ligand concentration -  1.0 nM!                           Positive Control:                                                                           Naloxone                                                        Incubation Conditions:                                                                      Naloxone                                                                      Reactions are carried out in 50 mM TRIS-HCl (pH 74) at                        25° C. for                                                             90 minutes. The reaction is terminated by rapid vacuum                        filtration                                                                    onto glass fiber filters. Radioactivity trapped onto the                      filters is                                                                    determined and compared to control values in order to                         ascertain                                                                     any interactions of test compound with the delta opiate                       binding site                                                                  Literature references to receptor binding activity                            procedure:                                                                    Akiyama, K., Gee, K. W., Mosberg, K. W. Yamamura, H. I.                       Characterization of  .sup.3 H!DPDPE Binding to Delta Opiate                   Receptors                                                                     in the Rat Brain and Neuroblastomaglioma Hybrid Cell Line                     (NG                                                                           108-115). Proc. Nat'l Acad. Sci. 82: 2543 (1985) with                         modifications.                                                                Sofuoglu, M., Portoghese, P. S., and Takemori, A. E.                          δ-Opioid                                                                Receptor Binding in Mouse Brain: Evidence for                                 Heterogeneous                                                                 Binding Sites. Eur. Jrnl. Pharm. 216: 273-277 (1992).           Results:      Receptor Binding Activity Percent Inhibition Average (N =                     2)                                                                            Nicotine Cotinine                                                                            Cotinine                                                                           Cotinine.sub.2 Fumarate                                   10.sup.-6 M                                                                            10.sup.-4 M                                                                         10.sup.-6 M                                                                        10.sup.-6 M                                               10.1%    10.0% 8.3% 17.6%                                       __________________________________________________________________________    Example 11.                                                                   Benzodiazepine (Peripheral) Binding Assay                                     __________________________________________________________________________    Reference Compounds Ki(nM)                                                                  PK11195                                                                            28.8                                                                     RO54864                                                                            34.0                                                                     Diazepam                                                                           574.0                                                      Assay Characteristics                                                         K.sub.d (binding affinity):                                                                 9.8 nM                                                          B.sub.max (receptor number):                                                                23.9 fmol/mg tissue (wet weight)                                Degree of Specific Binding:                                                                 90% (Non-specific binding determined using 10 μM                           PK11195)                                                        Materials and Methods:                                                        Receptor Source:                                                                            Rat kidney membranes                                            Radioligand:   .sup.3 H!PK11195 (60-90 Ci/mmol)                                             Final ligand concentration -  1.0 nM!                           Reference Compound:                                                                         PK11195                                                         Positive Control:                                                                           PK11195                                                         Incubation Conditions:                                                                      Reactions are carried out in 50 mM TRIS-HCl (pH 7.7) at                       0-4° C. for                                                            60 minutes. The reaction is terminated by rapid vacuum                        filtration                                                                    onto glass fiber filters. Radioactivity trapped onto the                      filters is                                                                    determined and compared to control values in order to                         ascertain                                                                     any interactions of test compound with the benzodiazepine                     (peripheral) binding site.                                                    Literature references to receptor binding activity                            procedure:                                                                    Skowronski, R., et al. Photoaffinity Labeling of Peripheral                   Type                                                                          Benzodiazepine Receptors in Rat Kidney Mitochondria with                       .sup.3 H!PK14105. Eur. Jrnl. Pharmac. 148: 187-193 (1988)                    with                                                                          modifications.                                                                Raghavendra Roa, V. L., Audet, R., Therrien, G., et al.                       Tissue                                                                        Specific Alterations of Binding Sites for Peripheral Type                     Benzodiazepine Receptor Ligand  .sup.3 H!PK11195 in Rats                      Following                                                                     Potacaval Anastomosis. Digestive Diseases & Sciences.                         39(5):                                                                        1055-1063 (1994).                                               Results:      Receptor Binding Activity Percent Inhibition Average (N =                     2)                                                                            Nicotine Cotinine                                                                            Cotinine                                                                           Cotinine.sub.2 Fumarate                                   10.sup.-6 M                                                                            10.sup.-4 M                                                                         10.sup.-6 M                                                                        10.sup.-6 M                                               3.1%     1.4%  8.5% 1.8%                                        __________________________________________________________________________    Example 12.                                                                   Glycine (Strychnine Insensitive) Binding Assay                                __________________________________________________________________________    Reference Compounds Ki(nM)                                                                  Glycine                                                                            187.0                                                                    D-Serine                                                                           703.0                                                                    D-Alanine                                                                          895.0                                                                    L-Serine                                                                           25,000                                                                   β-Alanine                                                                     900,000                                                    Assay Characteristics                                                         K.sub.d (binding affinity):                                                                 443 nM                                                          B.sub.max (receptor number):                                                                1.72 pmol/mg protein                                            Degree of Specific Binding:                                                                 90% (Non-specific binding determine using 1.0 mM glycine)       Materials and Methods:                                                        Receptor Source:                                                                            Rat cortical membranes                                          Radioligand:   .sup.3 H!Glycine (70-80 Ci/mmol)                                             Final ligand concentration -  10.0 nM!                          Reference Compound:                                                                         Glycine                                                         Positive Control:                                                                           Glycine                                                         Incubation Conditions:                                                                      Reactions are carried out in 50 mM HEPES (pH 7.1) at                          4° C. for 30                                                           minutes. The reaction is terminated by rapid vacuum                           filtration onto                                                               glass fiber filters. Radioactivity trapped onto the filters                   is                                                                            determined and compared to control values in order to                         ascertain                                                                     any interactions of test compound with the glycine binding                    site.                                                                         Literature references to receptor binding activity                            procedure:                                                                    Snell, L. D., Morter, R. S., and Johnson, K. M. Structural                    Requirements for Activation of the Glycine Receptor that                      Modulates                                                                     the N-methyl-D-aspartate Operated Channel. Eur. Jrnl.                         Pharmacol.                                                                    156: 105-110 (1988).                                                          Monahan, J. B., Corpus, V. M., Hood, W. F., et al.                            Characterization                                                              of a  .sup.3 H!Glycine Recognition Site as a Modulatory                       Site of the                                                                   NMDA Receptor Complex. Jrnl. Neurochem. 53(2): 370-375                        (1989).                                                         Results:      Receptor Binding Activity Percent Inhibition Average (N =                     2)                                                                            Nicotine Cotinine                                                                            Cotinine                                                                           Cotinine.sub.2 Fumarate                                   10.sup.-6 M                                                                            10.sup.-4 M                                                                         10.sup.-6 M                                                                        10.sup.-6 M                                               1.5%     -0.6% 10.9%                                                                              -5.6%                                       __________________________________________________________________________    Example 13.                                                                   Muscarinic - M.sub.1 Clone Binding Assay                                      __________________________________________________________________________    Reference Compounds Ki(nM)                                                                  Methylscopolamine                                                                        0.7                                                                4-DAMP-Methiodide                                                                        0.6                                                                p-F-Hexahydrodifenidol                                                                   8.5                                                                3-α-chloroimpenaline                                                               15.3                                                               Pirezepine 21.7                                                               Tropicamide                                                                              54.9                                                               Methoctramine                                                                            412.0                                                Assay Characteristics                                                         K.sub.d (binding affinity):                                                                 0.17 nM                                                         B.sub.max (receptor number):                                                                1,825 pmol/mg protein                                           Degree of Specific Binding:                                                                 90% (Non-specific binding determine using 1.0 μM                           Methylscopolamine bromide)                                      Materials and Methods:                                                        Receptor Source:                                                                            Human recombinant M.sub.1 clone                                 Radioligand:   .sup.3 H!Methylscopolamine (80-100 Ci/mmol)                                  Firial ligand concentration -  0.2 nM!                          Reference Compound:                                                                         (-)-Scopolamine, Methyl-, bromide (Methylscopolamine                          bromide)                                                        Positive Control:                                                                           (-)-Scopotamine, Methyl-, bromide (Methylscopolamine                          bromide)                                                        Incubation Conditions:                                                                      Reactions are carried out in 50 mM TRIS-HCl (pH 7.4)                          containing                                                                    10 nM MgCl.sub.2, 1 mM EDTA for 60 minutes at 25° C.                   The reaction is                                                               terminated by rapid vacuum filtration onto glass fiber                        filters.                                                                      Radioactivity trapped onto the filters is determined and                      compared                                                                      to control values in order to ascentain any interactions of                   test                                                                          compound(s) with the cloned muscarinic - M.sub.1 binding                      site.                                                                         Literature references to receptor binding activity                            procedure:                                                                    Wei, H. B., Roeske, W. R., et al. Pharmacological                             Characterization                                                              of a Novel Muscarinic Partial Agonist, YM796, in                              Transfected Cells                                                             Experssing the M.sub.1 or M.sub.2 Muscarinic Receptor Gene.                   Life Sciences.                                                                50(5): 355-363 (1992).                                                        Dorje, R., Wess, J., et al. Antagonist Binding Profiles of                    Five                                                                          Cloned Human Muscarinic Receptor Subtypes. Jrnl.                              Pharmacol.                                                                    Exp. Ther. 256(2): 727-733 (1991).                              Results:      Receptor Binding Activity Percent Inhibition Average (N =                     2)                                                                            Nicotine Cotinine                                                                            Cotinine                                                                           Cotinine.sub.2 Fumarate                                   10.sup.-6 M                                                                            10.sup.-4 M                                                                         10.sup.-6 M                                                                        10.sup.-6 M                                               0.9%     19.5% 15.5%                                                                              3.1%                                        __________________________________________________________________________    Example 14.                                                                   Muscarinic - M.sub.2 Clone Binding Assay                                      __________________________________________________________________________    Reference Compounds Ki(nM)                                                                  Methylscopolamine                                                                        0.5                                                                3-α-chloroimperialine                                                              0.7                                                                4-DAMP-Methiodide                                                                        10.1                                                               Tropicamide                                                                              19.4                                                               Methoctramine                                                                            58.8                                                               p-F-Hexahydrodifenidol                                                                   123.0                                                              Pirezepine 551.0                                                Assay Characteristics                                                         K.sub.d (binding affinity):                                                                 0.142 nM                                                        B.sub.max (receptor number):                                                                3,670 pmol/mg protein                                           Degree of Specific Binding:                                                                 90% (Non-specific binding determine using 1.0 μM                           Methylscopolamine bromide)                                      Materials and Methods:                                                        Receptor Source:                                                                            Human recombinant M.sub.2 clone                                 Radioligand:   .sup.3 H!Methylscopolamine (80-100 Ci/mmol)                                  Final ligand concentration -  0.2 nM!                           Reference Compound:                                                                         (-)-Scopotamine, Methyl-, bromide (Methylscopolamine                          bromide)                                                        Positive Control:                                                                           (-)-Scopolamine, Methyl-, bromide (Methylscopolamine                          bromide)                                                        Incubation Conditions:                                                                      Reactions are carried out in 50 mM TRIS-HCl (pH 7.4)                          containing                                                                    10 nM MgCl.sub.2, 1 mM EDTA for 60 minutes at 25° C.                   The reaction is                                                               terminated by rapid vacuum filtration onto glass fiber                        filters.                                                                      Radioactivity trapped onto the filters is determined and                      compared                                                                      to control values in order to ascertain any interactions of                   test                                                                          compound(s) with the cloned muscarinic - M.sub.2 binding                      site.                                                                         Literature references to receptor binding activity                            procedure:                                                                    Wei, H. B., Roeske, W. R., et al. Pharmacological                             Characterization                                                              of a Novel Muscarinic Partial Agonist, YM796, in                              Transfected Cells                                                             Experssing the M.sub.1 or M.sub.2 Muscarinic Receptor Gene.                   Life Sciences.                                                                50(5): 355-363 (1992).                                                        Dorje, R., Wess, J., et al. Antagonist Binding Profiles of                    Five                                                                          Cloned Human Muscarinic Receptor Subtypes. Jrnl.                              Pharmacol.                                                                    Exp. Ther. 256(2): 727-733(1991).                               Results:      Receptor Binding Activity Percent Inhibition Average (N =                     2)                                                                            Nicotine Cotinine                                                                            Cotinine                                                                           Cotinine.sub.2 Fumarate                                   10.sup.-6 M                                                                            10.sup.-4 M                                                                         10.sup.-6 M                                                                        10.sup.-6 M                                               4.4%     21.6% 22.3%                                                                              20.7%                                       __________________________________________________________________________    Example 15.                                                                   Serotonin - 5HT.sub.7 Clone Binding Assay                                     __________________________________________________________________________    Reference Compounds Ki(nM)                                                                  5-CT  0.6                                                                     Mesulergine                                                                         3.9                                                       Assay Characteristics                                                         K.sub.d (binding affinity):                                                                 3.2 nM                                                          B.sub.max (receptor number):                                                                710 fmol/mg protein                                             Degree of Specific Binding:                                                                 85% (Non-specific binding determine using 1.0 μM 5-                        carboxamidotryptamine, 5-CT)                                    Materials and Methods:                                                        Receptor Source:                                                                            recombinant Rat 5HT.sub.7 Clone                                 Radioligand:   .sup.3 H!LSD (60-80 Ci/mmol)                                                 Final ligand concentration -  2.0 nM!                           Reference Compound:                                                                         5-CT                                                            Positive Control:                                                                           5-CT                                                            Incubation Conditions:                                                                      Reactions are carried out in 50 mM TRIS-HCl (pH 7.4)                          containing                                                                    10 mM MgCl.sub.2, 1 mM EDTA for 60 minutes at 37° C.                   The reaction is                                                               terminated by rapid vacuum filtration onto glass fiber                        fiters.                                                                       Radioactivity trapped onto the filters is determined and                      compared                                                                      to control values in order to ascertain any interactions of                   test                                                                          compound(s) with the cloned serotonin - 5HT.sub.7 binding                     site.                                                                         Literature references to receptor binding activity                            procedure:                                                                    Shen, Y., et al. Molecular Cloning and Expression of a                        5-hydroxy-                                                                    tryptamine.sub.7 Serotonin Receptor Subtype. Jrnl. Biol.                      Chem. (268):                                                                  18200-18204 (1993).                                             Results:      Receptor Binding Activity Percent Inhibition Average (N =                     2)                                                                            Nicotine Cotinine                                                                            Cotinine                                                                           Cotinine.sub.2 Fumarate                                   10.sup.-6 M                                                                            10.sup.-4 M                                                                         10.sup.-6 M                                                                        10.sup.-6 M                                               8.0%     6.4%  -0.3%                                                                              1.4%                                        __________________________________________________________________________    Example 16.                                                                   Inositol Triphosphate (IP.sub.3) Binding Assay                                __________________________________________________________________________    Reference Compounds Ki(nM)                                                                  IP.sub.3                                                                         21                                                                         IP.sub.4                                                                         672                                                                        IP.sub.5                                                                         6,850                                                                      IP.sub.2                                                                         >10,000                                                                    IP.sub.1                                                                         >100,000                                                     Assay Characteristics                                                         K.sub.d (binding affinity):                                                                 40.0 nM                                                         B.sub.max (receptor number):                                                                23 pmol/mg protein                                              Degree of Specific Binding:                                                                 85% (Non-specific binding determined using 1.0 μM                          myo-inositol                                                                  1,4,5-triphosphate)                                             Materials and Methods:                                                        Receptor Source:                                                                            Rat cerebellar membranes                                        Radioligand:   .sup.3 H!IP.sub.3 (10-30 Ci/mmol)                                            Final ligand concentration -  4.0 nM!                           Reference Compound:                                                                         D-myo-inositol 1,4,5-triphosphate                               Positive Control:                                                                           D-myo-inositol 1,4,5-triphosphate                               Incubation Conditions:                                                                      Reactions are carried out in 50 mM TRIS-HCl (pH 8.3)                          containing                                                                    1 mM EDTA at 0° C. for 10 minutes. The reaction is                     terminated by                                                                 rapid vacuum filtration onto glass fiber filters.                             Radioactivity trapped                                                         onto the filters is determined and compared to control                        values in                                                                     order to ascertain any interactions of test compound with                     the                                                                           inositol triphospate binding site.                                            Literature references to receptor binding activity                            procedure:                                                                    Worley, P., Baraban, J., Supattapone, S., Wilson, V., and                     Snyder,                                                                       S. H. Characterization of Inositol Triphosphate Receptor                      Binding in                                                                    Brain. Jrnl. Biochem. 262(25): 12132-12136 (1987) with                        modifications.                                                                Willcocks, A. L., Cooke, A. M., Potter, B. V. L., and                         Nahorski, S. R.                                                               Stereospecific Recognition Sites for  .sup.3 H!Inositol (1,                   4, 5)-                                                                        Triphosphate in Particulate Preparations of Rat                               Cerebellum.                                                                   Biochem. Biophys. Res. Comm. 147: 1071-1078 (1987).             Results:      Receptor Binding Activity Percent Inhibition Average (N =                     2)                                                                            Nicotine Cotinine                                                                            Cotinine                                                                           Cotinine.sub.2 Fumarate                                   10.sup.-6 M                                                                            10.sup.-4 M                                                                         10.sup.-6 M                                                                        10.sup.-6 M                                               3.2%     8.3%  4.8% -5.1%                                       __________________________________________________________________________

In order to provide perspective concerning blood levels of the testcompounds employed, the molar concentrations listed above will berelated to cigarette consumption. For nicotine, a concentration of 10⁻⁶M equals a blood level of 166 ng/ml, which is a very high smoking doseof about 4-5 packs per day. For cotinine, 10⁻⁶ M equals a blood level of176 ng/ml, or a low smoking dose of about 1/4-1/2 pack per day. Cotinineat 10⁻⁴ M equals a blood level of 17,600 ng/ml, which is regarded to bean acceptable therapeutic dose level. Cotinine is so nontoxic that evenhigher doses can be used for short periods of time.

It has been shown previously that cotinine is capable of passing theblood-brain barrier (U.S. Pat. Nos. 3,870,794 and 5,187,169). Theresults obtained in the above examples show that the cotinine compoundsare, in addition, capable of binding to receptor sites. Cotinine andcotinine fumarate produced neuroleptic activity paralleling nicotine atall of the sites and neurotransmitters tested. In addition, the percentof inhibition for cotinine was numerically greater in 12 of the 16 itemsinvestigated. Cotinine therefore usually exhibits greater activity thannicotine at the same dosage. Moreover, since cotinine exhibits virtuallyno toxicity, it can be used therapeutically at much higherconcentrations than nicotine and therefore therapy with cotinine canproduce an effect many times greater than therapy with nicotine. In allof the examples presented, cotinine and cotinine fumarate exhibitbinding activity that is similar to nicotine and usually of greatermagnitude. The significance of this is that the examples thereforedemonstrate the similarity of cotinine and cotinine fumarate to nicotinein brain receptor binding assays at the binding sites and transmittersthat are important in Tourette's syndrome, obsessive-compulsive behaviorand schizophrenia. It is interesting to note that while neither nicotinenor cotinine bind especially to dopamine or serotonin receptors, bothcompounds are recognized to have dopaminergic and serotonergic activity.Presumably, these effects are pre-synaptic, post-synaptic or indirect.It should also be noted that the neurotransmitters in Examples 10, 11,13 and 14 are commonly implicated with drugs of abuse. These examplestherefore demonstrate therapeutic activity of cotinine in drug abuse.

The examples show that cotinine and cotinine fumarate have a bindingactivity that is as great as or greater than nicotine, and becausecotinine can be used therapeutically in a far higher concentration inthe blood, it can provide a much greater therapeutic effect thannicotine. The data from the above examples therefore provides supportfor cotinine compounds as a treatment for obsessive-compulsive disorder,Tourette's Syndrome and schizophrenia that is more effective thannicotine.

The Effect of Cotinine in Glutamatergic Transmission

McGehee previously reported that the behavioral and cognitive effects ofnicotine suggest that nicotinic acetylcholine receptors participate incentral nervous system (CNS) function. The complex behavioral effects ofnicotine are revealed by the finding that central nervous systemnicotinic acetylcholine receptors enhance fast excitatory transmission.In vitro studies also showed that one excitatory CNS transmitter isglutamate. This neurotransmitter is metabolized by the enzyme glutamicacid decarboxylase (GAD). The application of nicotine to the same CNSsites was shown by in vitro studies to enhance glutamatergictransmission since the amplitude of the evoked currents more thandoubled McGehee, et al. "Nicotine Enhancement of Fast ExcitatorySynaptic Transmission in CNS by Pre-synaptic Receptors", Science,269:1692 (1995)!. Enzyme activity studies conducted by me now show thatboth cotinine and nicotine have a similar effect on GAD as shown in thefollowing example.

Radioimmunoassay studies were conducted with rat corpus striatalmembranes using the following method.

    __________________________________________________________________________    Example 17.                                                                   Glutamic Acid Decarboxylase (GAD) Enzyme Assay                                __________________________________________________________________________    Reference Compounds Ki(nM)                                                                  (Amino-oxy)Acetic Acid                                                                   3,550                                                Assay Characteristics                                                         Degree of Specific Binding:                                                                 >95% (Non-specific activity determined in the absence of                      tissue)                                                         Materials and Methods:                                                        Enzyme Source:                                                                              Rat corpus striatal membranes                                   Substrate:     .sup.14 C!L-Glutamic acid (30-80 Ci/mmol)                      Reference Compound:                                                                         (Amino-oxy)Acetic Acid                                          Positive Control:                                                                           (Amino-oxy)Acetic Acid                                          Incubation Conditions:                                                                      Reactions are incubated in phosphate/mercapoethanol buffer                    (pH                                                                           6.9) for 30 minutes at 37° C. The reaction is                          stopped by the addition                                                       of trichloroacetic acid and placement on ice. Radioactive                     product is                                                                    trapped in phenlylethylamine/methanol and quantitated using                   liquid                                                                        scintillation chromatography in order to ascertain any                        interactions                                                                  of test compounds with the glutamic acid decarboxylase                        enzyme.                                                                       Literature references to receptor binding activity                            procedure:                                                                    Wilson, S. H., et al. Jrnl. Biol. Chem. 247: 3159 (1972).       Results:      Percent Inhibition                                                            Nicotine                                                                           Cotinine                                                                 10.sup.-8 M                                                                        10.sup.-6 M                                                              40%  38%                                                        __________________________________________________________________________

Example 17 shows that cotinine has about the same inhibitory effect ofnicotine with respect to GAD, an enzyme which converts glutamate intogammaaminobutyric acid (GABA). The ratio of glutamic acid to GABA(gammaaminobutyric acid) in the brain plays a crucial role in brainactivity. Glutamate is an excitatory transmitter. GABA is an inhibitorytransmitter. Furthermore, the concentration of glutamate and GABA arehigher in the brain than they are in the rest of the body. Theeffectiveness of cotinine to inhibit the metabolic enzyme GAD providesfurther support for its benefit in treating substance abuse or addictionby providing a "feeling good" response which lessens the need tocontinue the undesired activity or thought pattern. In treating thesedisorders, cotinine is administered in the same manner outlined above inany desired dosage form as already described. Because cotinine isthought to be innocuous when used by humans, people may be able to usecotinine as long as necessary in an effort to help them treat theirparticular addiction (or behavior) which may have developed over manyyears. This sheds light on the effectiveness of cotinine in treatingobsessive-compulsive disorder, Tourette's Syndrome and schizophreniathrough its ability to inhibit the activity of GAD so as to make thepatient feel better, thereby further assisting in alleviating orreducing the symptoms of the disease. Thus, nicotine and cotinine bothtrigger a molecular switch in the brain that speeds up the flow andintensity of glutamate, a function important in the nicotine-dependencyprocess because it elicits a "feeling-good" response in the person whohas consumed the nicotine, but cotinine will produce a therapeuticeffect many times greater than nicotine because it can be safelyadministered in much greater amounts.

The present invention has utility in treating other psychiatricdisorders. The data developed in the above examples indicate that theinvention is useful in substance addiction generally and, in particular,in treating drug and alcohol addiction so as to alleviate or reduce thesymptoms of the disorder and in treating manic-depression syndrome byproviding a "feeling good" response when administered as described asabove. The invention is also indicated to be useful in alleviating orreducing the symptoms of anorexia and bulimia, which are characterizedby a failure to eat appropriate amounts of food or by eating andpurging. In these disorders it is the same nicotinic-acetylcholinereceptors, as well as others described in the above examples, which areactive in the chain of events that current research indicates to be animportant factor underlying the disease. The examples demonstrate thatcotinine is effective in influencing these receptors and transmitters inan advantageous manner so as to relieve or reduce the symptoms of thedisorder. The heavy use of tobacco by drug and alcohol addicts furthersupports the effectiveness of cotinine in these applications because itis the main metabolite of the nicotine that was consumed. Moreover,anorexia and bulimia are indeed forms of an obsessive-compulsivedisorder which are indicated to respond favorably to treatment using thepresent invention as already noted. Accordingly, bulimia/anorexiapatients can benefit from the present invention in a manner similar topatients who suffer from other forms of obsessive-compulsive disorders.It should be noted that behavioral scientists group obsessive-compulsivedisorder, Tourette's Syndrome, and anorexia/bulimia together because oftheir similar neurochemical origins.

Nicotine has been described as a "behavioral homeostat" because itappears to increase "low behavior" and decrease "high behavior"Rosecrans J. A. "The Role of Nicotine in Tobacco Dependence andCessation." Chemistry and Industry, pp.221-224 (21 Mar. 1994)!. Theauthor explains the mechanism by noting "nicotine's ability todesensitize its receptor." The results of the tests described aboveindicate that the cotinine metabolized from nicotine, which acts toblock the nicotinic cholinergic receptors (in effect desensitizingthem), and the dynamic interplay of varying concentrations of nicotineand cotinine in the smoker's brain produces the unusual benefitpreviously described as a "behavioral homeostat."

All publications and patents are herein incorporated by reference to thesame extent as if each individual publication or patent was specificallyand individually indicated to be incorporated by reference.

Many variations of the present invention within the scope of theappended claims will be apparent to those skilled in the art once theprinciples described herein are understood.

What is claimed is:
 1. A method for treating a patient suffering fromobsessive-compulsive disorder, the method comprising,administering tothe patient an effective amount of cotinine or a pharmaceuticallyacceptable salt thereof, which amount is effective to reduce oralleviate at least one of the symptoms of obsessive-compulsive disorderin a human or other mammal.
 2. The method of claim 1 wherein theeffective amount of the cotinine or salt thereof is from about 0.5 mg/kgto about 100 mg/kg of body weight per day.
 3. The method of claim 1wherein the cotinine is (-)-cotinine.
 4. The method of claim 1 whereinthe cotinine is a salt of (-)-cotinine.
 5. The method of claim 1 whereinthe cotinine or salt is a pharmaceutical unit dosage form selected froma unit dosage for oral administration, a peritoneal administrationdosage form, a transdermal delivery form, an interocular administrationdosage form, an intravenous administration dosage form, a dosage formwhich comprises cotinine or the cotinine salt dissolved in a liquidvehicle, an intranasal administration dosage form, an inhalation dosageform.
 6. The method of claim 5 wherein the cotinine or thepharmaceutically acceptable salt thereof is administered in a tablet orcapsule.
 7. A method for treating a patient suffering from Tourette'sSyndrome, the method comprising,administering to the patient aneffective amount of cotinine or a pharmaceutically acceptable saltthereof, which amount is effective to reduce or alleviate at least oneof the symptoms of Tourette's Syndrome in a human or other mammal. 8.The method of claim 7 wherein the effective amount of the cotinine orsalt thereof is from about 0.5 mg/kg to about 100 mg/kg of body weightper day.
 9. The method of claim 7 wherein the cotinine is (-)-cotinine.10. The method of claim 7 wherein the cotinine is a salt of(-)-cotinine.
 11. The method of claim 7 wherein the cotinine or salt isa pharmaceutical unit dosage form selected from a unit dosage for oraladministration, a peritoneal administration dosage form, a transdermaldelivery form, an interocular administration dosage form, an intravenousadministration dosage form, a dosage form which comprises cotinine orthe cotinine salt dissolved in a liquid vehicle, an intranasaladministration dosage form, an inhalation dosage form.
 12. The method ofclaim 11 wherein the cotinine or the pharmaceutically acceptable saltthereof is administered in a tablet or capsule.
 13. A method fortreating a patient suffering from schizophrenia, the methodcomprising,administering to the patient an effective amount of cotinineor a pharmaceutically acceptable salt thereof, which amount is effectiveto reduce or alleviate at least one of the symptoms of schizophreniadisorder in a human or other mammal.
 14. The method of claim 13 whereinthe effective amount of the cotinine or salt thereof is from about 0.5mg/kg to about 100 mg/kg of body weight per day.
 15. The method of claim13 wherein the cotinine is (-)-cotinine.
 16. The method of claim 13wherein the cotinine is a salt of (-)-cotinine.
 17. The method of claim13 wherein the cotinine or salt is a pharmaceutically unit dosage formselected from a unit dosage for oral administration, a peritonealadministration dosage form, a transdermal delivery form, an interocularadministration dosage form, an intravenous administration dosage form, adosage form which comprises cotinine or the cotinine salt dissolved in aliquid vehicle, an intranasal administration dosage form, an inhalationdosage form.
 18. The method of claim 17 wherein the cotinine or thepharmaceutically acceptable salt thereof is administered in a tablet orcapsule.
 19. The method of claim 13 wherein the administration ofcotinine is accompanied by the administration of haloperidol to thepatient.